CLONING AND CHARACTERIZATION OF A NEW PROTEASE GENE (PRTH) FROM PORPHYROMONAS-GINGIVALIS

Porphyromonas gingivalis has been implicated as a contributing etiolog ical agent of adult periodontitis and generalized forms of early-onset periodontitis. Proteases of P. gingivalis may contribute to its patho genicity by destroying connective tissue as well as inactivating keg p lasma proteins that might mediate protective host functions. In order to explore this problem, antiserum raised against membrane vesicles of P. gingivalis W83 was used to screen a genomic library of strain W83 constructed by using the lambda DASH vector system. A recombinant phag e (lambda 34) expressing a P. gingivalis protease from the library was identified and characterized. Casein substrate zymography of lambda 3 4 lysates revealed a protease with an apparent molecular mass of 97 kD a. The gene encoding this protease was designated prtH. It was localiz ed to a 3.7-kb HindIII-BamHI fragment and specified an enzyme which hy drolyzed the human C3 complement protein under defined conditions. The nucleotide sequence of this 3.7-kb fragment was determined, and one 2 .9-kb open reading frame (992 amino acids) corresponding to a 110-kDa protein was detected, suggesting it might be a precursor of the 97-kDa active protease. prtH is not similar to any previously cloned proteas e gene from P. gingivalis.